Below, the main steps are discussed and the optimised steps in respect to the published literature are highlighted.
The results are compared to a non-optimised step in acceleration and to a nonlinear closed loop control of the air path.
Of the five one-step kits tested using the Mx3005P cycler, three optimised one-step kits (i.e., Quantitect-1 step, Rotor Gene-1 step, and SSIII Platinium-1 step) revealed good sensitivity with the detection of at least 10 copies/ µL.
The height of each riser was optimised in Step II using a detailed FE model with the optimised riser positions from Step I.
For all optimised one-step or two-step PCR assays, the interassay CVs were lower than 10%, demonstrating good reproducibility for all tested kits [ 23].
Electrode design, optimisation steps and characteristics for the optimised electrode configuration are presented.
Whenever regression models are optimised, it is important that all optimisation steps are properly validated.
The two other optimised one-step assays (i.e., Verso-1 step and qScript-1 step) revealed poor sensitivity with a detection of 10 and 10 copies/ µL, respectively.
We found that the optimised one-step PCR assays had a higher detection sensitivity than the optimised two-step assays regardless of the machine used, while no difference was detected in reaction efficiency, R values, and intra- and interreproducibility between the two methods.
In our experience, regardless of the real-time cycler used, one-step qRT-PCR showed better limit of detection at 95% than the optimised two-step assays using pan Lyssavirus primers.
The strong cube texture was obtained through an optimised two-step recrystallisation annealing following heavy cold working.
